BACKGROUND: Smad7 is a major repressible protein in transforming growth factor β(TGF-β1) signal transduction pathway,which possess antifibrotic effects.OBJECTIVE: To construct rat Smad7 eukaryotic vector and to observe the mRNA expression level of TGF-β1, collagen Ⅰ and collagen Ⅲ in rat hepatic stellate cells (HSC)-T6 cell.DESIGN, TIME AND SETTING: The gene recombination and cytology observation experiment was performed at the First Affiliated Hospital of Shihezi University School of Medicine.MATERIALS: pcDNA3.1 (+) plasmid was reserved in the laboratory. E coil DH5α was presented by Key Laboratory of Xinjiang Endemic and Ethnic Diseases, Shihezi University School of Medicine. The HSC T6 cell was provided by Cancer Institute and Hospital, Chinese Academy of Medical Sciences.METHODS: Rat Smad7 cDNA was cloned into eukaryotic plasmid pcDNA3.1(+) to construct Smad7/pcDNA3.1(+) plasmid and transfected it into HSC-T6 ceils by Lipofectmine 2000. The experiment was divided into normal control, empty vector and Smad7 transfected groups, and the positive cells were selected by G418.MAIN OUTCOME MEASURES: The levels of Smad7, TGF-β1, collagen Ⅰ and Ⅲ mRNA was detected by reverse transcriptase polymerase chain reaction, respectively.RESULTS: Smad7 eukaryotic vector was successfully constructed and confirmed by endonuilease digestion and sequencing. Compared to the control and empty vector groups, Smad7 mRNA expression was significantly higher in Smad7 transfected group (P < 0.01 ); and TGF-β1 and collagen Ⅰ mRNA expression was notably reduced (P < 0.01). There was no statistically significant difference of the change of collagen Ⅲ mRNA expression among the three groups (P>0.05). The difference of Smad7, TGF-β1,collagen Ⅰ and Ⅲ mRNA expression had no statistically significant between control and empty vector groups (P_(all) > 0.05)CONCLUSION: Smad7 eukaryotic expression vector is successfully constructed. The Smad7 gene can effectively expressed in transfected HSC-T6 cell, and decrease mRNA expressions of TGF-β1 and collagen Ⅰ.

The effect of phenol concentration on the structure and function of microbial communities,which were cultured in different conditions using coking wastewater biofilm as seeding,was investigated by Biolog and denaturing gradient gel electrophoresis(DGGE)methods.The less number of bands of cultivated sam-ples on the denaturing gradient gel electrophoresis fingerprint of 16S rRNA gene indicated reduction of di-versity after enrichment and cultivation.Some bands on the DGGE gel were significantly influenced by the phenol concentration in medium.The results of Biolog showed that the original biofilm sample had the highest substrate utility capacity as measured by average well color development(AWCD).But low concen-tration of phenol enriched sample S7 showed more diverse activity on the utility of Carboxylic acids.The principal component analysis(PCA)of Biolog data revealed that the metabolic patterns were similar when using the same phenol concentration,although the sample S7 much less similar to other cultivated samples.These results suggested that the enrichment and cultivation with phenol supplemented decreased the diver-sity and also changed the metabolic function of the microbial community.Lower phenol concentration in-creased the microbial community metabolic activity.The phenol degrading capacity of isolates from each samples indicated that the enrichment and cultivation condition had changed the type and property of cul-truable bacteria.Based on these results,we concluded that the different microorganisms will be isolated un-der different cultivation condition.

Objective To study the relationship between activity of thorax and each spinal intervertebral angle in patients with ankylosing spondylitis. Methods Each spinal intervertebral angle of 41 patients with ankylosing spondylitis were measured by Spinalmouse in different postures. And the activity of thorax was measured. Correlation between activity of thorax and shape of spinal were analyzed. Results The activity of thorax positively correlated with the entire lumbar spinal column in flexion (P<0.01), as well as the intervertebral angle of L1/2, L2/3 and L4/5 in flexion (P<0.05), but negatively correlated with the intervertebral angle of L1/2 and L2/3, curvature of the entire lumbar spinal column in upright and the intervertebral angle of L1/2, L3/4, curvature of the entire lumbar spinal column in extension. Conclusion There was a significant relation between activity of thorax and lumbar vertebra motor ability.

Five compounds (tenuifoliside C, tenuifoliside D, telephiose A, telephiose C and polygalaxanthone III) from polygala tenuifolia wild were incubated together with CYP probe substrate in human liver microsomes to investigate the inhibitory effect towards CYP450 enzyme. Phenacetin (CYP1A2), coumarin (CYP2A6), paclitaxel (CYP2C8), diclofenac (CYP2C9), S-mepheriytoin (CYP2C19), dextromethorphan (CYP2D6), chlorzoxazone (CYP2E1), midazolam (CYP3A) were selected as the isoforfn specific substrate. And the formation of paracetamol, 7-hydroxycoumarin, 6alpha-hydroxy paclitaxel, 4'-hydroxydiclofenac, dextrorphan, 6-hydroxychlorzoxazone, 1'-hydroxymidazolam, 4'-hydroxymephenytoin were detected respectively to measure the effect towards CYP450 by high-pressure liquid chromatography (HPLC). The result shows that five compounds from polygala tenuifolia willd significantly inhibit chlorzoxazone 6-hydroxylation catalyzed by CYP2E1, while showed no effect towards CYP1A2, CYP2A6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP3A. And IC50 value was 38.73, 54.14, 61.77, 62.22, 50.56 micromol x L(-1), respectively.
Cytochrome P-450 Enzyme System ; metabolism ; Esters ; pharmacology ; Glycosides ; pharmacology ; Humans ; Microsomes, Liver ; drug effects ; enzymology ; Oligosaccharides ; pharmacology ; Polygala ; chemistry ; Xanthones ; pharmacology

To study the compatible mechanisms and compatible proportion of Shaoyao Gancao decoction, the intestinal absorption of main ingredients in Shaoyao Gancao decoction SG11 (Baishao-Zhigancao 1: 1) , SG31 (Baishao-Zhigancao 3: 1), Baishao water decoction S and Zhigancao (G) were investigated and compared using in vitro everted intestinal sac model and in situ single pass intestinal perfusion (SPIP) model. The concentration of paeoniflorin (PF), liquiritin (LQ) and mono-ammonium glycyrrhizinate (GL) in test samples and samples of intestinal sac and intestinal perfusion was determined by HPLC. The intestinal absorptive amount and absorption parameters were calculated. Results showed that in the everted intestinal sac model, three ingredients could be absorbed by duodenum, jejunum and ileum, and the absorption in the jejunum was best for all 3 ingredients. The absorption rate of three ingredients in SG11 was significantly higher than that in single decoction (P < 0.05), but had no significant difference compared with SG31. In SPIP model, the absorption rate constant K(a), the apparent absorption coefficient P(app) and the absorption rate of three ingredients in SG11 were significantly higher than those in single decoction. Parameters of PF and GL in SG11 were significantly higher than those in SG31, but had no differences of LQ. It proved that the compatibility of Baishao and Zhigancao could improve the intestinal absorption of PF, LQ and GL. The absorption of each ingredient in SG11 was better than that in SG31.
Animals ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; pharmacokinetics ; Intestinal Absorption ; drug effects ; Intestines ; blood supply ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley

Through the analysis of the problems of health service delivery system in rural areas and the causes of the abnormal operation of the three-tier rural health service network, this article probed into the continuity of health service in rural China. These issues are observed from the aspects of continuity of disciplines, institutions, relationship and health information. Policy recommendations include remolding the institutional relations in the three-tier rural health service network, constructing reasonable supporting environment and improving the health delivery quality.

Objective To analyze the expression characteristics of SRPK2 ( serine /arginine-rich protein specific kinase 2 mRNA and its encoded protein products in mouse testis, and to reveal its molecular role during spermatogenesis. Methods Using semi-quantitative reverse transcription polymerase chain reaction ( RT-PCR) and Western blotting to an-alyze the expression of SRPK2 mRNA and its protein products in several mouse tissues.The stage-specific expression pat-tern of SRPK2 mRNA in mouse testis was examined by real-time quantitative PCR.Immunohistochemical staining was ap-plied to identify the SRPK2 protein distribution in mouse testicular seminiferous tubules and indirect immunofluorescence staining was used to detect the subcellular localization of SRPK2 protein in spermatic cells.Results Semi-quantitative RT-PCR and Western blotting analysis revealed that SRPK2 was highly expressed in mice testis.Real-time quantitative PCR analysis showed that SRPK2 mRNA was expressed abundantly at the stage of 5-and 8-week old mouse testes, suggesting that SRPK2 gene has stage-specific expression patterns during mouse spermatogenesis.Immunohistochemical and indirect immu-nofluorescence staining demonstrated that SRPK2 protein was located around the surface of elongated sperm nucleus. Conclusions SRPK2 is highly expressed in mouse testis and has significant stage-specific expression characteristics with distinct nuclear localization.These results indicate that SRPK2 may participate in precursor mRNA splicing during mouse spermiogenesis.The molecular mechanism of SRPK2 is yet to be further studied.

Adolescent ; Child ; Child, Preschool ; Complement System Proteins ; analysis ; Female ; Humans ; Immunoglobulins ; analysis ; Infant ; Male ; Pneumonia, Mycoplasma ; immunology

Objective:To investigate the effect of dihydromyricetin (DMY) on LPS-induced septic shock in mice and the related underlying mechanism.Methods:The LPS-induced septic shock mice model was established after the mice were pre-treated by DMY for 7 days.The mortality rate was calculated at 24,48,72,96,120,144 and 168 h after the mice were intraperitoneal injected with LPS.For elucidation of underlying mechanism ,RAW246.7 were pre-incubated with DMY for 1 h,and then stimulated by LPS 100 ng/ml.Western blot was performed for determination of P-ERK,P-JNK and P-p38 expression.Immunohistochemistry was applied to explore c-Fos and c-Jun nucleus translocation.Results:DMY could significantly inhibit LPS-induced mice mortality.Inhibitory effect of DMY on the phosphorylation of JNK and p 38 contributed to the anti-inflammatory effect of DMY in vivo.Furthermore , DMY obviously prevented c-Fos and c-Jun nucleus translocation.Conclusion:The anti-inflammatory effect of DMY is attributed to the suppression on c-Fos and c-Jun nucleus translocation ,via inhibition of the phosphorylation of JNK and p 38.

Objective To investigate the mechanism of tumor necrosis factor-α (TNF)-α inhibiting osteo blastdifferentiation of mesenchymal stem cells (BMMSCs) in the pathogenesis of osteoporosis in the mouse model of systemic lupus erythematosus (MRL/lpr). Methods The femurs of MRL / lpr and C3He/HeJ mice were isolated, the bone structure were examined by hematoxylin-eosin (HE) staining. The proteins of TNF-α, NF-κB P50, bone morphogenetic protein -2 (BMP-2) and PSmad1/5/8 were measured by immunohistochemical stain. Bone marrow mesenchymal stem cells (BMMSCs) were isolated. After BMMSCs grew on the cover slips, the proteins on top of it were evaluated by immunohistochemistry stain. Moreover, the alkaline phosphatase (ALP) staining was employed for the measurement of the early osteogenic differentiation. BMMSCs together with hydroxyapatite were embedded subcutaneously in the nude mice and eight weeks later, the ectopic bone formation was evaluated. The recombinant human tumor necrosis factor receptor type Ⅱantibody fusion protein (etanercept) or normal saline was subcutaneous injected to the mice with lupus. After four weeks, the expression of these proteins was observed and the ectopic bone formation was investigated. Image-Pro plus 6.0 software was employed for imagine analysis, and Studentˊs t-test was used to test the differences between 2 independent groups. Results MRL/lpr mice showed decreased volume of cortex and the percentage of cortex to the volume of bone of MRL/lpr mice was significantly lower compared to control groups and with C3He/HeJ mice (13.96±0.25 vs 23.61±0.71, n=3, P<0.01). The protein levels of both TNF-αand NF-κB P50 on the femur of MRL/lprl mice were higher than those of the control group (0.643±0.051 vs 0.405±0.022, 0.917±0.023 vs 0.650±0.032, n=3, P<0.01). The expressions of BMP-2 on the femur of MRL/lpr mice were lower than those of the C3He/HeJ mice (0.52 ±0.03 vs 0.72 ±0.03, n=3, P<0.01). There was no difference in the expression of PSmad1/5/8 on the femur between the two groups by immunohistochemistry detection (1.264 ±0.021 vs 1.301± 0.044, n=3, P>0.05). The expressions of TNF-α and NF-κB P50 in BMMSCs of MRL/lprl mice were higher than those of the C3He/HeJ (0.184±0.021 vs 0.136±0.013, 0.132±0.021 vs 0.097± 0.014, n=3, P<0.01), while BMP-2 and PSmad were lower than those of the control group (0.128±0.013 vs 0.216±0.221, 0.115±0.023 vs 0.196±0.034, n=3, P<0.01). After 7 days of BMP-2 stimulation, the activities of ALP of BMMSCs from MRL/lprl mice were reduced detected by ALP staining and the osteoblast differentiation of these cells were decreased than BMMSCs from the control mice by HE and Masson staining. The percentage of the cortex to the volume of bone of the etanercept injection MRL/lpr mice was higher than that of the control group (21.8±1.0 vs 14.3 ±0.6, n=3, P<0.01). Moreover, the proteins of TNF-α and NF-κB P50 on the femurs of such injected mice were lower than those of the control group (0.540±0.024 vs 0.682±0.031, 0.857±0.023 vs 1.098±0.044, n=3, P<0.05), while the expressions of BMP-2 were higher than the control group (0.99±0.04 vs 0.85±0.04, n=3, P<0.05). There was no difference in the PSmad1/5/8 expression on the bone of the two group of lupus mice (0.88 ±0.08 vs 0.84 ±0.04, n=3, P>0.05). The ectopic bone formation of BMMSCs of the etanercept injected MRL/lpr mice was higher than that of the normal saline injected mice, however, it was lower than that of the C3He/HeJ mice. Conclusion TNF-α inhibits osteoblast differentiation of mesenchymal stem cells by depressing Smad signaling which may contribute to the osteoporosis of the lupus mice.